Impact of variant histology on upstaging and survival in patients with nonmuscle invasive bladder cancer undergoing radical cystectomy.

INTRODUCTION: Variant histology (VH) of urothelial carcinoma is uncommon and frequently presents at the muscle-invasive stage. VH is considering a significant risk factor for progression among patients with nonmuscle invasive bladder cancer (NMIBC). While there is some debate, expert opinion is generally that upfront radical cystectomy (RC) should be consider for these patients. Limited data exists to support this position. In this study, we sought to examine the rate of upstaging and overall survival for patients with VH NMIBC against patients with pure urothelial NMIBC who underwent RC, to help clarify the optimal treatment strategy for these patients. METHODS: The institutional REDCap database was utilized to identify all patients with T1 and Ta bladder cancer that underwent RC over the study period (2004-2022). Matched-pair analysis was performed between patients with VH and pure urothelial NMIBC; 42 pairs were matched on prior intravesical therapy, presence of muscularis propria on transurethral resection of bladder tumor (TURBT), any carcinoma in situ presence on prior TURBTs, and final tumor staging on TURBT. The primary outcomes of interest were pathologic tumor upstaging rate at RC and overall survival. Secondary outcomes of interest included association of demographic or pretreatment variables with upstaging, and upstaging rates for specific variant histologies. RESULTS: Patients with VH NMIBC undergoing RC were upstaged at a significantly higher rate than a matched cohort of patients with pure urothelial NMIBC (73.8% vs. 52.4%, P = 0.0244) and among those upstaged, had significantly higher rates of pT3 to pT4 (54.7% vs. 23.8%, P = 0.0088). Rate of node positivity at RC for VH NMIBC was also higher compared to pure urothelial NMIBC (40.5% vs. 21.4%, P = 0.0389). Among histologic variants, patients with plasmacytoid and sarcomatoid subtypes demonstrated the highest rates of upstaging; differences were not statistically significant. The overall median survival was 28.4 months for patients with VH after RC compared to 155.1 months for patients with pure urothelial NMIBC (P = 0.009). CONCLUSION: Patients with VH NMIBC undergoing RC are at significantly higher risk of upstaging at RC when compared to patients with pure urothelial NMIBC and have worse overall survival. While this study supports the concept of an aggressive treatment approach for patients with VH NMIBC, improvements in understanding of the disease are necessary to improve outcomes.

Role of transmembrane 4 superfamily (TM4SF) proteins CD9 and CD81 in muscle cell fusion and myotube maintenance.

The role of transmembrane 4 superfamily (TM4SF) proteins during muscle cell fusion has not been investigated previously. Here we show that the appearance of TM4SF protein, CD9, and the formation of CD9-beta1 integrin complexes were both regulated in coordination with murine C2C12 myoblast cell differentiation. Also, anti-CD9 and anti-CD81 monoclonal antibodies substantially inhibited and delayed conversion of C2C12 cells to elongated myotubes, without affecting muscle-specific protein expression. Studies of the human myoblast-derived RD sarcoma cell line further demonstrated that TM4SF proteins have a role during muscle cell fusion. Ectopic expression of CD9 caused a four- to eightfold increase in RD cell syncytia formation, whereas anti-CD9 and anti-CD81 antibodies markedly delayed RD syncytia formation. Finally, anti-CD9 and anti-CD81 monoclonal antibodies triggered apoptotic degeneration of C2C12 cell myotubes after they were formed. In summary, TM4SF proteins such as CD9 and CD81 appear to promote muscle cell fusion and support myotube maintenance.

Overexpression of the TGFbeta-regulated zinc finger encoding gene, TIEG, induces apoptosis in pancreatic epithelial cells.

Members of the TGFbeta family of peptides exert antiproliferative effects and induce apoptosis in epithelial cell populations. In the exocrine pancreas, these peptides not only regulate normal cell growth, but alterations in these pathways have been associated with neoplastic transformation. Therefore, the identification of molecules that regulate exocrine pancreatic cell proliferation and apoptotic cell death in response to TGFbeta peptides is necessary for a better understanding of normal morphogenesis as well as carcinogenesis of the pancreas. In this study, we have characterized the expression and function in exocrine pancreatic epithelial cells of the TGFbeta-inducible early gene (TIEG), a Krüppel-like zinc finger transcription factor encoding gene previously isolated from mesodermally derived osteoblastic cells. We demonstrate that this gene is expressed in both acinar and ductular epithelial cell populations from the exocrine pancreas. In addition, we show that the expression of TIEG is regulated by TGFbeta1 as an early response gene in pancreatic epithelial cell lines. Moreover, overexpression of TIEG in the TGFbeta-sensitive epithelial cell line PANC1 is sufficient to induce apoptosis. Together, these results support a role for TIEG in linking TGFbeta-mediated signaling cascades to the regulation of pancreatic epithelial cell growth.

PTEN mutation, EGFR amplification, and outcome in patients with anaplastic astrocytoma and glioblastoma multiforme.

BACKGROUND: Survival of patients with anaplastic astrocytoma is highly variable. Prognostic markers would thus be useful to identify clinical subsets of such patients. Because specific genetic alterations have been associated with glioblastoma, we investigated whether similar genetic alterations could be detected in patients with anaplastic astrocytoma and used to identify those with particularly aggressive disease. METHODS: Tissue specimens were collected from 174 patients enrolled in Mayo Clinic Cancer Center and North Central Cancer Treatment Group clinical trials for newly diagnosed gliomas, including 63 with anaplastic astrocytoma and 111 with glioblastoma multiforme. Alterations of the EGFR, PTEN, and p53 genes and of chromosomes 7 and 10 were examined by fluorescence in situ hybridization, semiquantitative polymerase chain reaction, and DNA sequencing. All statistical tests were two-sided. RESULTS: Mutation of PTEN, amplification of EGFR, and loss of the q arm of chromosome 10 were statistically significantly less common in anaplastic astrocytoma than in glioblastoma multiforme (P =.033, P =.001, and P<.001, respectively), and mutation of p53 was statistically significantly more common (P<.001). Univariate survival analyses of patients with anaplastic astrocytoma identified PTEN (P =.002) and p53 (P =.012) mutations as statistically significantly associated with reduced and prolonged survival, respectively. Multivariate Cox analysis of patients with anaplastic astrocytoma showed that PTEN mutation remained a powerful prognostic factor after adjusting for patient age, on-study performance score, and extent of tumor resection (hazard ratio = 4.34; 95% confidence interval = 1.82 to 10.34). Multivariate classification and regression-tree analysis of all 174 patients identified EGFR amplification as an independent predictor of prolonged survival in patients with glioblastoma multiforme who were older than 60 years of age. CONCLUSION: PTEN mutation and EGFR amplification are important prognostic factors in patients with anaplastic astrocytoma and in older patients with glioblastoma multiforme, respectively.

Gene therapy for carcinoembryonic antigen-producing human lung cancer cells by cell type-specific expression of herpes simplex virus thymidine kinase gene.

A carcinoembryonic antigen (CEA)-producing human lung cancer cell line (A549), a nonproducing human lung cancer cell line (CADO-LC9), and a human uterine cervical cancer (HeLa) were transfected with the herpes simplex virus thymidine kinase (HSV-TK) gene regulated by 445 nucleotides upstream from the translational start of CEA gene. Fifty % growth inhibitory concentration of ganciclovir (GCV) was 0.57 micron for HSV-TK-transfected A549; relative sensitivity to GCV was more than 1000 times higher compared to the 50% growth inhibitory concentration of the parental cell line. Both CADO-LC9 and HeLa transfected with HSV-TK were still resistant to GCV. There was no difference in either morphology or doubling time between HSV-TK-transfected and parental clones. Injections (i.p.) of GCV resulted in significant regression of HSV-TK-transfected A549 tumors in nude mice. These data show the possibility of gene therapy using the cell type-specific promoter of CEA gene against CEA-producing adenocarcinoma of the lung.

Eradication of Myc-overexpressing small cell lung cancer cells transfected with herpes simplex virus thymidine kinase gene containing Myc-Max response elements.

Herpes simplex virus thymidine kinase (HSV-TK) gene was ligated with four repeats of the Myc-Max response elements (a core nucleotide sequence CACGTG), and its utility for gene therapy was examined by the treatment of either c-, L- or N-myc-overexpressing the small cell lung cancer (SCLC) cell line with ganciclovir (GCV). The chloramphenicol acetyltransferase assay demonstrated that the overexpression of any myc genes activated transcription from the CAT gene depending on the Myc-Max binding sites. The transduction of the HSV-TK gene ligated with the CACGTG core rendered all three SCLC lines to be more sensitive to GCV than parental ones in vitro. In addition, the growth of c- or L-myc-overexpressing SCLC cells containing the hybrid HSV-TK gene were significantly suppressed by GCV in vivo. When parental SCLC cells were mixed with HSV-TK-expressing tumor cells at a ratio of 1:3, GCV treatment inhibited tumor growth by 90% compared with parental cells only, indicating the existence of the "bystander effect." These data suggest that the CACGTG-driven HSV-TK gene may be useful for the treatment of SCLC overexpressing any type of myc family oncogenes.

A frequent deletion of chromosome 5q21 in advanced small cell and non-small cell carcinoma of the lung.

We have examined the deletion of the long arm of chromosome 5 (5q) in 59 cases of advanced lung cancer [39 cases of small cell lung cancer (SCLC), 20 cases of non-SCLC] using 12 restriction fragment length polymorphism markers on 5q. Of 59 lung cancer cases, 48 (81%) exhibited deletion at any portion of the 5q locus (loci). Such a high frequency of 5q deletion has not been reported in surgically resectable non-SCLC. One SCLC case showed a 5q deletion only in metastatic sites but not in the primary cancer. These data suggest that the inactivation of putative tumor-suppressor gene(s) on 5q may be a late event in the progression of lung cancer. There was no significant difference in frequency of 5q deletion between SCLC and non-SCLC. Compared to non-SCLC, however, SCLC usually showed widespread deletion on 5q. While the most frequent target region was estimated to be about 3-5 megabases at 5q21 around the adenomatous polyposis coli (APC) gene locus, some cases showed more telomeric deletion (5q33-35), suggesting that there are at least two different tumor-suppressor genes on 5q associated with the progression of lung cancer.

Application of the Cre recombinase/loxP system further enhances antitumor effects in cell type-specific gene therapy against carcinoembryonic antigen-producing cancer.

A considerable number of studies of cancer have shown that the cell type-specific promoter is an effective tool for selective expression of foreign genes in tumor cells. However, few reports have demonstrated significant in vivo antitumor effects using this strategy thus far, possibly because the low activity of such a promoter results in insufficient expression of genes in cancer cells as well as in insignificant antitumor effects, even when the cells are infected by highly efficient gene transfer methods. To overcome this problem, we used the Cre/loxP system for the cell type-specific gene therapy against carcinoembryonic antigen (CEA)-producing cancer. We constructed a pair of recombinant Ads. One expresses the Cre recombinase (Cre) gene under the control of the CEA promoter (Ad.CEA-Cre). The other contains the herpes simplex virus thymidine kinase (HSV-TK) gene separated from the strong CAG promoter by insertion of the neomycin resistance (neo) gene (Ad.lox-TK). The HSV-TK gene of the latter Ad is designed to be activated through excisional deletion of the neo gene by Cre enzyme released from the former one only when CEA-producing cells are infected simultaneously with these Ads. Coinfection by these Ads rendered a human CEA-producing cancer cell line 8.4-fold more sensitive to ganciclovir (GCV) compared with infection by Ad.CEA-TK alone, the HSV-TK gene of which is directly regulated by the CEA promoter. On the other hand, coinfection with these Ads did not significantly change the GCV sensitivity of non-CEA-producing cells. Intratumoral injection of Ad.CEA-Cre combined with Ad.lox-TK followed by GCV treatment almost completely eradicated CEA-producing tumors established in the subcutis of athymic mice, whereas intratumoral injection of Ad.CEA-TK with GCV administration at most retarded the growth of inoculated tumors. These results suggest distinct advantages of the Cre/loxP system applied in the conventional cell type-specific gene therapy against cancer.

Generation of a small cell lung cancer variant resistant to lymphokine-activated killer (LAK) cells: association with resistance to a LAK cell-derived, cytostatic factor.

Cells of OS2-RA, a human small cell lung cancer line sensitive to lymphokine-activated killer (LAK) cells, were repeatedly cocultured with human LAK cells. Fourteen cycles of the coculture produced a variant, termed OS2-RA-R, capable of growing successfully in the presence of LAK cells. OS2-RA-R showed a moderate resistance to lysis by LAK cells in 4-h 51Cr release assays. OS2-RA-R acted positively as a cold target for lysis of OS2-RA by LAK cells, suggesting no loss of the binding site for LAK cells on the cell surface of the variant. On the other hand, LAK cells were shown to produce a factor capable of suppressing the proliferation of OS2-RA and certain other cell lines but not lymphocytes. Interestingly, OS2-RA-R exhibited a substantial resistance to the cytostatic activity of LAK cell supernatants. The cytostatic factor, eluted at the 57-kDa fraction in gel filtration, showed no activity of interleukin 1, gamma-interferon, transforming growth factor beta, or tumor necrosis factor. These results suggest that LAK cells exhibit antitumor activity through not only rapid cytolysis but also slow-acting cytokine production, and the successful growth of OS2-RA-R in a coculture with LAK cells is the result of acquiring resistance to these two different LAK cell phenomena.

Pharmacological profile of TP-680, a new cholecystokininA receptor antagonist.

1. The pharmacological characteristics of a newly developed serine derivative (R)-1-[3-(3-carboxypyridine-2-yl) thio-2-(indol-2-yl)carbonylamino]propionyl-4-diphenylmethyl- piperazine (TP-680), a cholecystokinin type A (CCKA) receptor antagonist, were studied and compared with those of MK-329 and loxiglumide. 2. TP-680 showed approximately 2 and 22 times greater selectivity for peripheral CCKA receptors relative to brain CCK (CCKB) receptors than MK-329 and loxiglumide, respectively, when IC50 values for inhibition of [125I]-CCK-8 binding in isolated acini and cerebral cortex were compared. 3. TP-680 was approximately 17 times less potent than MK-329, but was 106 times more potent than loxiglumide in inhibiting 100 pM CCK-8-stimulated amylase release from rat pancreatic acini. The antagonism produced by TP-680 was specific for CCK in that the effects of other receptor secretagogues or agents bypassing receptors were not altered. 4. TP-680 caused a parallel rightward shift of the dose-response curve for CCK-8-stimulated amylase release as did MK-329 and loxiglumide. However, in contrast to MK-329 and loxiglumide, TP-680 suppressed the maximal responses of CCK-8-induced amylase release in a concentration-dependent fashion, indicating that TP-680 is an unsurmountable antagonist. 5. Repeated washing of acini after a 30 min treatment with TP-680 restored the responsiveness but not the sensitivity, causing a residual inhibition on the action of CCK-8. 6. The addition of loxiglumide prior to or together with application of TP-680 protected CCK receptors from unsurmountable and irreversible antagonism by TP-680. 7. Our results indicate that TP-680 is a potent and the most selective CCKA receptor antagonist for the pancreas reported to date.

Investigation of germline PTEN, p53, p16(INK4A)/p14(ARF), and CDK4 alterations in familial glioma.

Epidemiological studies suggest that some familial aggregations of glioma may be due to inherited predisposition. Many genes involved in familial cancers are frequently altered in the corresponding sporadic forms. We have investigated several genes known to be altered in sporadic gliomas for their potential contribution to familial glioma. Fifteen glioma patients with a family history of brain tumors were identified through the Mayo Clinic Department of Neurology (nine diffuse astrocytomas, two oligodendrogliomas, two mixed oligoastrocytomas, one pilocytic astrocytoma, and one pineal glioma). Eleven of the propositi had one or more first degree relative with a glioma. Lymphocyte DNA was derived from each of the patients and analyzed by polymerase chain reaction (PCR) and direct sequencing of the PTEN, p53, p16(INK4A)/p14(ARF), and CDK4 genes. In addition, fluorescence in situ hybridization (FISH) was performed on EBV-transformed lymphocytes from each affected individual to detect germline copy number of the p16(INK4A)/p14(ARF) tumor suppressor region. A p53 germline point mutation was identified in one family with some findings of Li-Fraumeni syndrome, and a hemizygous germline deletion of the p16(INK4A)/p14(ARF) tumor suppressor region was demonstrated by FISH in a family with history of both astrocytoma and melanoma. Thus, whereas germ-line mutations of PTEN, p53, p16(INK4A)/p14(ARF), and CDK4 are not common events in familial glioma, outside of familial cancer syndromes, point mutations of p53 and hemizygous deletions and other rearrangements of the p16(INK4A)/p14(ARF) tumor suppressor region may account for a subset of familial glioma cases. Collectively, these data lend genetic support to the heritable nature of some cases of glioma.

Elevated levels of soluble interleukin-2 receptors in tuberculous pleural effusions.

The levels of soluble IL-2R were measured in pleural fluid from patients with tuberculosis pleurisy. There were significantly elevated soluble IL-2R values in tuberculous pleural fluid as compared with pleural fluid of nontuberculous etiology including malignant, bacterial and transudative pleural effusions. In patients with tuberculous pleurisy, the level of soluble IL-2R in pleural fluid was markedly greater than that in serum. Furthermore, a significant positive correlation was observed between soluble IL-2R levels and adenosine deaminase levels in tuberculous pleural fluid. These findings suggest that elevated levels of pleural fluid soluble IL-2R in tuberculous pleurisy could reflect the local proliferation of activated T-cells and may be clinically useful in the diagnostic procedures for patients with pleural tuberculosis.

Complementary DNA sequence encoding the major neural cell adhesion molecule isoform in a human small cell lung cancer cell line.

The neural cell adhesion molecule (N-CAM), a member of the immunoglobulin gene super-family mediating homophilic cell-cell adhesion in a neuroendocrine system, is preferentially expressed in human small cell lung cancer (SCLC). Immunoprecipitation of a panel of SCLC cell lines by monoclonal antibodies (mAbs) specific for N-CAM detects mainly the 145-kDa isoform. This result was correlated with Northern blotting where a single 6.2-kb mRNA was detected in nine SCLC cell lines. To determine cDNA sequence encoding the N-CAM isoform, we selected several cDNA clones encoding N-CAM isolated from OS2-R, a SCLC cell line established in our laboratory. Based on the analysis of the full-length cDNA obtained from two clones, the sequence of this 145-kDa isoform was shown to be essentially identical to that of the 140-kDa N-CAM isoform of neuroblastoma except for a single base pair changed at position 1620 without changing amino acid encoded.

Detailed deletion mapping of the short arm of chromosome 3 in small cell and non-small cell carcinoma of the lung.

We constructed a detailed deletion map of the short arm of chromosome 3 (3p) for 55 lung cancer cases by using 17 restriction fragment length polymorphism (RFLP) probes. Initially, we examined 40 small cell lung cancer (SCLC) cases and found three regions of deletion at 3p25-26, 3p21.3 and 3p14-cen, suggesting the possibility of at least three different tumor-suppressor genes on 3p. In order to obtain more detailed deletion area, and to compare the pattern of 3p deletion, we also examined 15 non-small cell lung cancer (NSCLC) cases. Compared to NSCLC cases, most of SCLC cases have widespread deletion on 3p, suggesting multiple tumor-suppressor genes on 3p may be inactivated in this type of cancer. In 3p21.3 area, minimum overlapping area of deletion lays between two probes which are close to each other. These data will be useful to isolate the putative tumor-suppressor genes located on the chromosome 3p.

Treatment with cholecystokinin receptor antagonist loxiglumide enhances insulin response to intravenous glucose stimulation in postpancreatitic rats.

Pancreatic exocrine and endocrine function in postpancreatitic rats treated with cholecystokinin (CCK) receptor antagonist loxiglumide was compared with that treated with saline and CCK octapeptide (CCK-8) or with that in normal control rats. Treatment with loxiglumide (50 mg/kg body weight), CCK-8 (2.5 micrograms/kg body weight), or saline (2.5 ml/kg body weight) was given three times a day for 6 days starting 1 day after the induction of acute pancreatitis by a 4-h subcutaneous infusion of 20 micrograms/kg body weight/h of caerulein. On day 8, pancreatic exocrine and endocrine function was simultaneously determined following an intravenous injection of a mixed solution of 0.2 g/kg body weight glucose plus 100 ng/kg body weight caerulein. Basal pancreatic juice flow was significantly increased in all of the postpancreatitic rats irrespective of the treatment, whereas the maximal juice flow in the loxiglumide- and saline-treated rats was significantly low compared with the CCK-8-treated and the control rats. Basal and the peak protein outputs in the loxiglumide-treated rats were comparable to those in saline-treated rats, but were lower than those in the control or the CCK-8-treated rats. Although serum glucose concentrations in all of the postpancreatitic rats were similar to those in the control rats, stimulated as well as basal insulin release tended to be high compared with the control rats. In particular, loxiglumide-treated rats showed the exaggerated insulin response compared with other groups of rats. These present observations indicate that administration of high dose of loxiglumide for a long period decreases pancreatic enzyme output and causes insulin resistance.

Effects of cholecystokinin receptor antagonist loxiglumide on rat exocrine pancreas.

Effects of long-term administration of the cholecystokinin receptor antagonist loxiglumide on exocrine pancreas were studied in adult rats. Plasma concentrations of loxiglumide at 8 h after a single subcutaneous injection of 50 mg/kg body weight of loxiglumide were 3.2 +/- 0.8 microgram/ml, which were comparable to those at 12 h after oral administration of the same dose (3.7 +/- 0.9 microgram/ml). Eight hours' prior subcutaneous injection of loxiglumide (50 mg/kg body weight) significantly suppressed pancreatic exocrine secretion stimulated by an intravenous bolus injection of 50 ng/kg body weight caerulein compared with the control rats. Based on these results, in the first experiment, loxiglumide at a dose of 50 mg/kg body weight was given subcutaneously three times a day (low dose) for 6 days to adult rats fed a standard laboratory diet. Low dose of loxiglumide significantly decreased pancreatic wet weight (-14%) and pancreatic contents of protein (-26%), trypsin (-38%), and lipase (-68%), while having no significant effect on pancreatic contents of DNA and amylase. In the second experiment, three times higher dose of loxiglumide (150 mg/kg body weight) was given by an orogastric tube twice daily for 6 days. High dose of loxiglumide significantly decreased pancreatic weight (-11%) and contents of protein (-20%) and DNA (-22%), whereas it significantly increased amylase (+92%) and trypsin content (+20%).(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of the cholecystokinin receptor antagonist loxiglumide on pancreatic exocrine function in rats after acute pancreatitis.

Recent studies have demonstrated that cholecystokinin (CCK) receptor antagonists not only reduce the severity of pancreatitis but also inhibit pancreatic regeneration after pancreatitis. This study was undertaken, therefore, to examine the effects of the CCK receptor antagonist loxiglumide on the exocrine pancreas when given after an episode of acute pancreatitis that was induced in rats by a 4-h subcutaneous infusion of 20 micrograms/kg body weight/h cerulein. Biochemical changes and secretory function in response to 100 ng/kg body weight cerulein were determined after a 6-day treatment with saline, loxiglumide (50 mg/kg body weight), or CCK-8 (2.5 micrograms/kg body weight), which was given three times a day starting 24 h after the induction of acute pancreatitis. In the saline-treated rats, pancreatic enzyme contents and pancreatic juice and protein output were significantly low, whereas the pancreatic weight and protein and DNA contents were comparable to those of the controls without pancreatitis. Loxiglumide treatment, although significantly decreasing protein output, had no influence on pancreatic weight, protein and DNA contents, or pancreatic juice flow but increased the amylase and lipase contents compared to those of the saline-treated postpancreatitic rats. CCK-8 treatment also had no influence on pancreatic weight or protein and DNA contents but significantly increased the pancreatic enzyme contents and pancreatic juice and protein output compared to those of the saline-treated postpancreatitic rats. These results suggest that loxiglumide does not significantly inhibit the recovery of exocrine function but appears to accelerate the increase in pancreatic amylase and lipase contents even when given after an attack of acute pancreatitis.

Role of endogenous cholecystokinin and cholecystokinin-A receptors in the development of acute pancreatitis in rats.

Recent studies provide significant evidence that cholecystokinin (CCK) is involved in the induction and development of acute pancreatitis in experimental animals. However, the results obtained with specific CCK-A (peripheral) receptor antagonists are still controversial. The present studies were undertaken to evaluate the involvement of endogenous CCK and the CCK-A receptors in the development of severe acute pancreatitis induced in Otsuka Long-Evans Tokushima Fatty (OLETF) rats that have a selective defect in the CCK-A receptor. Three models of severe acute pancreatitis were induced by retrograde intraductal infusion of 4% sodium taurocholate, by the closed duodenal loop, or by a single intraperitoneal injection of 500 mg/100 g body weight of L-arginine in OLETF rats and control Long-Evans Tokushima Otsuka (LETO) rats. Plasma CCK levels rose up to 4- to 14-fold over the preloading values after the onset of acute pancreatitis in all three models in both groups of rats. However, histologic alterations as well as the magnitudes of increase in serum amylase and lipase activity and the pancreatic wet weight were significantly less in the OLETF rats than those in the LETO rats. In addition, 72 h after the onset of arginine pancreatitis, massive destruction of pancreatic parenchyma with a significant reduction in serum amylase and lipase activities and pancreatic wet weight was observed in the LETO rats, whereas these changes were not seen in OLETF rats. These results suggest that endogenous CCK and CCK-A receptors play a role in the development of severe acute pancreatitis in rats.

Ethanol inhibits CCK-induced enzyme secretion by affecting calcium-pump activity in isolated rat pancreatic acini.

The aim of this study was to clarify the effect of ethanol on stimulus-secretion coupling in pancreatic exocrine secretion. We investigated the effects of 600 mM ethanol on cholecystokinin octapeptide (CCK-8)-stimulated amylase release, cytosolic free Ca2+ concentration ([Ca2+]i) and Ca2+ fluxes using in vitro isolated rat pancreatic acini. Ethanol, given alone, stimulated both the initial and the sustained phases of amylase release. On the other hand, ethanol inhibited only the sustained phase of amylase release stimulated by CCK-8. Ethanol also inhibited amylase release in response to fluoride, a direct activator of guanine nucleotide-binding protein, suggesting that ethanol affects intracellular signal transduction molecules. Ethanol had no influences on the initial rise but increased the sustained rise in [Ca2+]i stimulated by CCK-8 and inhibited CCK-8-stimulated Ca2+ outflux without affecting Ca2+ influx. 8-Bromoguanosine 3':5'-cyclic monophosphate, a membrane-permeable analogue of cGMP regulating membrane Ca(2+)-pump activity in various cells, completely reversed the ethanol-induced inhibition of amylase release and Ca2+ outflux in response to CCK-8 as well as fluoride. Given that Ca2+ plays a critical role in stimulus-secretion coupling in pancreatic exocrine secretion, our results indicate that 600 mM ethanol inhibits CCK-8-stimulated amylase release by inhibiting Ca(2+)-pump activity on the plasma membrane.

Effects of MCI-727 on pancreatic exocrine secretion and acute pancreatitis in two experimental rat models.

The effects of a newly developed compound having antiulcer action, (Z)-2-(4-methylpiperazin-1-yl)-1-[4-(2-phenyl-ethyl)phenyl]-eth anone oxime hydrochloride monohydrate (MCI-727), on pancreatic exocrine secretion were studied in anesthetized rats and evaluated its preventive and therapeutic effects on acute pancreatitis in two experimental rat models. Intraduodenal administration of MCI-727 [25, 50, or 100 mg/kg body weight (wt)] stimulated a dose-dependent increase in pancreatic juice and bicarbonate output without increasing the protein output or plasma cholecystokinin concentration. MCI-727-stimulated pancreatic exocrine secretion was completely abolished by antisecretin serum but not by the cholecystokinin receptor antagonist loxiglumide (50 mg/kg body wt/h) or cholinergic receptor antagonist atropine (100 mu g/kg body wt/h). In rats with acute pancreatitis induced by four subcutaneous injections of 20 mu g/kg body wt cerulein at hourly intervals over 3 h, MCI-727 administered orally at a dose of 100 mg/kg body wt 30 min before the first cerulein injection significantly reduced the increases in serum amylase and lipase activity and pancreatic wet weight and induced improvements in the results of histologic examination. Moreover, when given 30 min before and 90 min after the first cerulein injection, MCI-727 had even more dramatic protective effects on all these parameters. In addition, even when administered immediately after the last cerulein injection, MCI-727 effectively ameliorated all these alterations of acute pancreatitis. However, MCI-727 had no apparent beneficial effects on the biochemical and histologic alterations of acute pancreatitis in the severe form induced by retrograde intraductal injection of 1.0 ml/kg body wt of 4% sodium taurocholate. These findings suggest that oral administration of MCI-727 stimulates pancreatic exocrine secretion by endogenous secretin release and that it has therapeutic as well as preventive effects on mild forms of acute pancreatitis in rats.